Fluorescent PCR (polymerase chain reaction) detection kit for vibrio cholerae and systematic identification method thereof

Abstract

The invention provides a quick detection method and a fluorescent PCR (polymerase chain reaction) detection kit invented according to the typical biochemical characteristics of sucrose, arabinose, mannose, oxidase, haircuts, salt-free peptone water and the like of the vibrio cholerae. The invention designs primers and TaqMan probes according to the outer-membrane protein gene (ompW), the toxin expression regulation protein gene (toxR) and the hemolysin gene (hlyA), wherein the vibrio cholerae outer-membrane protein gene is used as a characteristic gene for identifying the vibrio cholerae, andthe toxin expression regulation protein gene and the hemolysin gene are used for discriminating toxin genes carried by the vibrio cholerae. The fluorescent detection method for detecting the vibrio cholerae and related toxin genes thereof is provided based on the three genes. According to the detection method, strains are primarily screened through biochemical reaction, the strains with positive results are confirmed by fluorescent quantitative PCR, and the strains with positive ompW genes are vibrio cholerae. The vibrio cholerae and related toxin genes thereof can be quickly, accurately and specifically detected by the biochemical reaction and fluorescent quantitative PCR.

Description

(1) Technical field [0001] The invention relates to a fluorescent PCR detection kit for three groups of Vibrio cholerae and a method for systematic identification of three groups of Vibrio cholerae using the kit. (2) Background technology [0002] Cholera is a kind of severe infectious disease caused by Vibrio cholerae with diarrhea as the main symptom. It is one of the ancient and widespread severe infectious diseases and has caused many pandemics in the world. Vibrio cholerae is the causative agent of cholera in humans. Vibrio cholerae has been divided into more than 200 serogroups according to the different bacterial (O) antigens, of which only O1 and O139 groups can cause cholera pandemics, and the rest of the serotypes are widely distributed in natural water bodies, generally not disease or only sporadic cases of diarrhea and intestinal infections. [0003] The Diarrhea Control Center of the World Health Organization divides Vibrio cholerae into 3 groups: O1 group Vib...

AI Technical Summary

Problems solved by technology

The traditional methods for detection of Vibrio cholerae mainly include morphology, biochemistry, coagulation test, etc. The cycle of these detections is long, generally more than 18 to 24 hours. strains) cannot be detected

Although some immunological methods are fast, the results are unstable, and the current routine detection methods only detect Vibrio cholerae of O1 and O139 serotypes, and cannot detect non-O1 and non-O139 serotype strains

At the same time, studies have clarified that virulence genes such as toxR and hlyA can be transferred from cholera O1 group toxin-producing strains to environmental non-O1 and O139 group strains, enhancing the toxicity of non-O1 and O139 group strains. Studies have shown that non-O1 and non-O139 group Group Vibrio cholerae is more suitable for survival and reproduction in seawater products than O1 and O139 group Vibrio cholerae, so it greatly threatens the food safety of seawater products

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